It is useful for diagnosing or detecting a disease process to perform a histologic or cytologic examination of a tissue cell sample using a light microscope. This requires that a tissue (cellular material) sample must first be retrieved from the patient, and then processed for microscopic examination. A number of minimally invasive techniques are available for retrieving and collecting cell samples from a patient, e.g., by using a fine needle aspiration biopsy, or by brushing body cavity surfaces accessible through minimally invasive endoscopic techniques. A variety of cell sample processing techniques are also known, such as the Cytospin® technique and the Thin-prep® technique, for depositing cellular materials and tissue fragments directly onto a microscope slide. Another technique, commonly referred to as a cell block preparation, immobilizes cellular materials and/or small tissue fragments within a solid support structure, typically paraffin, to form a “cell block”. Thin sections of the cell block are then cut with a microtome and mounted onto a microscope slide for examination.
U.S. Pat. No. 6,913,921 (“the '921 patent”) discloses and describes methods and apparatus for cell block preparation, including providing a tissue collection cassette that serves a dual function of capturing cellular sample matter and providing a fluid pathway through which the cell processing and embedding reagents can flow. The cellular sample material is provided in an aqueous solution or a liquid cell preservative, which is passed through the tissue cassette across a filter that traps the cells and tissue fragments. A reagent flow pathway is configured to sequentially pass embedding reagents (alcohol, xylene, eosin stain) and liquefied paraffin through the tissue cassette and the cell sample already deposited on the filter. Once the paraffin is cooled, the filter is peeled away, leaving a paraffin “disk” protruding from the tissue cassette with embedded cellular matter positioned at the end of the disk in a plane at which a tissue section can be cut using a standard microtome for microscope examination. U.S. patent application Ser. No. 11/839,531, filed on Aug. 15, 2007, and assigned to the assignee of the present invention (“the '531 application”), discloses a substantially automated cell block creation that does not require human oversight during creation, including a two-piece cassette and filter assembly, to achieve more consistent cellular matter quantities in the created cell blocks, shorter processing time, reduced use of hazardous reagents, and more fully encapsulated cell blocks to preserve nucleic acid integrity. The contents of the respective '921 patent and '531 application are hereby fully incorporated by reference as if fully set forth herein.
When dispensing sample fluid into collection wells in the disclosed processes of the '531 application, certain samples will tend to clog the filter before a sufficient cellular material layer is retained. Such samples usually contain small individual cells, such as lymphocytes and other inflammatory cells, which have a tendency to stack and almost immediately impede the fluid flow across the filter. In order to collect additional cellular materials, and therefore a larger cell layer retained by the filter, air bubbles are pushed (or pulsed) through the filter from the waste chamber to temporarily lift the cellular materials away from the filter surface, and allow more liquid to pass through. This is done by applying a pressure within the waste chamber just larger than the bubble point of the filter and sample liquid combination to gently lift the cellular material from the filter surface. When sample collection has stalled, this small pressure pulse can lift the sample from the filter and allow more sample to be collected.
The bubble point is the pressure required on one side of a filter to produce bubbles on the other side of the filter, which is in contact with a liquid. The bubble point varies depending on the pore size of the filter and the surface tension of the liquid with which the filter is in contact. The bubble point of a filter can be manually calibrated by incrementally increasing the pressure applied to the non-liquid side of the filter (“back pressure”) and visually observing when bubbles first form. The back pressure at which bubbles first form is the bubble point of the filter and liquid combination.
In the disclosed processes of the '531 application, the bubble point of the filter and sample liquid combination is known to the user prior to initiating cell block creation. If the pressure applied is lower than the bubble point, the impeding cellular materials will not be sufficiently lifted away from the filter surface to allow for efficient sample processing. If the pressure applied is exceeds the bubble point by a large amount, the impeding cellular materials may be ejected from the sample well of the cassette. This may lead to loss of valuable sample and may also occlude equipment mounted above the sample well, such as an ultrasonic liquid level sensor, resulting in spurious level detection. Thus, while representing an improvement over the then-state of the art for cell block preparation, the methods and apparatuses disclosed in the '921 patent and the '531 application require using a pre-calibrated filter with a known bubble point to determine the appropriate back pressure to dislodge impeding cellular material without ejecting it from the sample well of a cassette.